The long-range objective of this project is to develop methods to modulate peptide hormone action through the peptidases that inactivate them. The focus of this application is to understand the contribution of three metallopeptidases, insulysin, neprilysin, neprilysin-2, and a novel plasma membrane (3-endorphin cleaving enzyme, to neuropeptide metabolism. The three specific aims are: 1. To study the allosteric properties of insulysin and its contribution to modulating physiological peptide levels. Insulysin, which hydrolyzes a variety of peptides including (3-endorphin, insulin, and amyloid (3 peptides is allosterically regulated by both substrates (homotrophic effects) and by anions such as ATP (heterotrophic effects). The heterotrophic effectors switch the preference of the enzyme from cleaving relative large peptides like insulin, amyloid (3-peptide, and -3-endorphin to smaller peptides such as the dynorphins. 2. To study the contribution gate the relative contributions of neprilysin and neprilysin-2 to neuropeptide metabolism in the brain. 3. To isolate and characterize a newly found plasma membrane peptidase that degrades (3-endorphin. We propose to characterize this novel peptidase, determine its tissue distribution as well as its substrate specificity. Relevence: Together these studies should contribute to our understanding of the involvement of a group of peptidases in the regulation of peptide action and may lead to new ways in which to modulate these activities in pathological states. For example one of the peptidases to be studied has recently been implicated in HIV dependent dementia in elderly affected individuals. 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